6at3: Difference between revisions
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==E. coli phosphoenolpyruvate carboxykinase Y207F mutant bound to thiosulfate and oxaloacetate== | |||
<StructureSection load='6at3' size='340' side='right'caption='[[6at3]], [[Resolution|resolution]] 1.46Å' scene=''> | |||
== Structural highlights == | |||
<table><tr><td colspan='2'>[[6at3]] is a 2 chain structure with sequence from [https://en.wikipedia.org/wiki/Escherichia_coli_K-12 Escherichia coli K-12]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=6AT3 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=6AT3 FirstGlance]. <br> | |||
</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 1.455Å</td></tr> | |||
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=OAA:OXALOACETATE+ION'>OAA</scene>, <scene name='pdbligand=THJ:THIOSULFATE'>THJ</scene></td></tr> | |||
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=6at3 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=6at3 OCA], [https://pdbe.org/6at3 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=6at3 RCSB], [https://www.ebi.ac.uk/pdbsum/6at3 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=6at3 ProSAT]</span></td></tr> | |||
</table> | |||
== Function == | |||
[https://www.uniprot.org/uniprot/PCKA_ECOLI PCKA_ECOLI] | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
Protein engineering to alter recognition underlying ligand binding and activity has enormous potential. Here, ligand binding for Escherichia coli phosphoenolpyruvate carboxykinase (PEPCK), which converts oxaloacetate into CO2 and phosphoenolpyruvate as the first committed step in gluconeogenesis, was engineered to accommodate alternative ligands as an exemplary system with structural information. From our identification of bicarbonate binding in the PEPCK active site at the supposed CO2 binding site, we probed binding of nonnative ligands with three oxygen atoms arranged to resemble the bicarbonate geometry. Crystal structures of PEPCK and point mutants with bound nonnative ligands thiosulfate and methanesulfonate along with strained ATP and reoriented oxaloacetate intermediates and unexpected bicarbonate were determined and analyzed. The mutations successfully altered the bound ligand position and orientation and its specificity: mutated PEPCKs bound either thiosulfate or methanesulfonate but never both. Computational calculations predicted a methanesulfonate binding mutant and revealed that release of the active site ordered solvent exerts a strong influence on ligand binding. Besides nonnative ligand binding, one mutant altered the Mn(2+) coordination sphere: instead of the canonical octahedral ligand arrangement, the mutant in question had an only five-coordinate arrangement. From this work, critical features of ligand binding, position, and metal ion cofactor geometry required for all downstream events can be engineered with small numbers of mutations to provide insights into fundamental underpinnings of protein-ligand recognition. Through structural and computational knowledge, the combination of designed and random mutations aids in the robust design of predetermined changes to ligand binding and activity to engineer protein function. | |||
Structural Control of Nonnative Ligand Binding in Engineered Mutants of Phosphoenolpyruvate Carboxykinase.,Tang HYH, Shin DS, Hura GL, Yang Y, Hu X, Lightstone FC, McGee MD, Padgett HS, Yannone SM, Tainer JA Biochemistry. 2018 Dec 4;57(48):6688-6700. doi: 10.1021/acs.biochem.8b00963. Epub, 2018 Nov 15. PMID:30376300<ref>PMID:30376300</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
[[Category: | </div> | ||
<div class="pdbe-citations 6at3" style="background-color:#fffaf0;"></div> | |||
==See Also== | |||
*[[Phosphoenolpyruvate carboxykinase 3D structures|Phosphoenolpyruvate carboxykinase 3D structures]] | |||
== References == | |||
<references/> | |||
__TOC__ | |||
</StructureSection> | |||
[[Category: Escherichia coli K-12]] | |||
[[Category: Large Structures]] | |||
[[Category: Shin DS]] | |||
[[Category: Tainer JA]] | |||
[[Category: Tang HYH]] | |||
Latest revision as of 17:24, 4 October 2023
E. coli phosphoenolpyruvate carboxykinase Y207F mutant bound to thiosulfate and oxaloacetateE. coli phosphoenolpyruvate carboxykinase Y207F mutant bound to thiosulfate and oxaloacetate
Structural highlights
FunctionPublication Abstract from PubMedProtein engineering to alter recognition underlying ligand binding and activity has enormous potential. Here, ligand binding for Escherichia coli phosphoenolpyruvate carboxykinase (PEPCK), which converts oxaloacetate into CO2 and phosphoenolpyruvate as the first committed step in gluconeogenesis, was engineered to accommodate alternative ligands as an exemplary system with structural information. From our identification of bicarbonate binding in the PEPCK active site at the supposed CO2 binding site, we probed binding of nonnative ligands with three oxygen atoms arranged to resemble the bicarbonate geometry. Crystal structures of PEPCK and point mutants with bound nonnative ligands thiosulfate and methanesulfonate along with strained ATP and reoriented oxaloacetate intermediates and unexpected bicarbonate were determined and analyzed. The mutations successfully altered the bound ligand position and orientation and its specificity: mutated PEPCKs bound either thiosulfate or methanesulfonate but never both. Computational calculations predicted a methanesulfonate binding mutant and revealed that release of the active site ordered solvent exerts a strong influence on ligand binding. Besides nonnative ligand binding, one mutant altered the Mn(2+) coordination sphere: instead of the canonical octahedral ligand arrangement, the mutant in question had an only five-coordinate arrangement. From this work, critical features of ligand binding, position, and metal ion cofactor geometry required for all downstream events can be engineered with small numbers of mutations to provide insights into fundamental underpinnings of protein-ligand recognition. Through structural and computational knowledge, the combination of designed and random mutations aids in the robust design of predetermined changes to ligand binding and activity to engineer protein function. Structural Control of Nonnative Ligand Binding in Engineered Mutants of Phosphoenolpyruvate Carboxykinase.,Tang HYH, Shin DS, Hura GL, Yang Y, Hu X, Lightstone FC, McGee MD, Padgett HS, Yannone SM, Tainer JA Biochemistry. 2018 Dec 4;57(48):6688-6700. doi: 10.1021/acs.biochem.8b00963. Epub, 2018 Nov 15. PMID:30376300[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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